The metabolism of diclofenac to its 5-hydroxylated derivative in humans is
catalyzed by cytochrome P450 (CYP)3A4. We report herein that in vitro this
biotransformation pathway is stimulated by quinidine. When diclofenac was i
ncubated with human liver microsomes in the presence of quinidine, the form
ation of 5-hydroxydiclofenac increased similar to 6-fold relative to contro
ls. Similar phenomena were observed with diastereoisomers of quinidine, inc
luding quinine and the threo epimers, which produced an enhancement in the
formation of 5-hydroxydiclofenac in the order of 6- to 9-fold. This stimula
tion of diclofenac metabolism was diminished when human liver microsomes we
re pretreated with a monoclonal inhibitory antibody against CYP3A4. In cont
rast, neither cytochrome b(5) nor CYP oxidoreductase appeared to mediate th
e stimulation of diclofenac metabolism by quinidine, suggesting that the ef
fect of quinidine is mediated through CYP3A4 protein. Further kinetic analy
ses indicated that V-max values for the conversion of diclofenac to its 5-h
ydroxy derivative increased 4.5-fold from 13.2 to 57.6 nmol/min/nmol of CYP
with little change in K-m (71-56 mu M) over a quinidine concentration rang
e of 0 to 30 mu M. Conversely, the metabolism of quinidine was not affected
by the presence of diclofenac; the K-m value estimated for the formation o
f 3-hydroxyquinidine was similar to 1.5 mu M, similar to the quinidine conc
entration required to produce 50% of the maximum stimulatory effect on dicl
ofenac metabolism. It appears that the enhancement of diclofenac metabolism
does not interfere with quinidine's access to the ferriheme-oxygen complex
, implicating the presence of both compounds in the active site of CYP3A4 a
t the same time. Finally, a similar to 4-fold increase in 5-hydroxydiclofen
ac formation was observed in human hepatocyte suspensions containing diclof
enac and quinidine, demonstrating that this type of drug-drug interaction o
ccurs in intact cells.